Polyethylene terephthalate, also known as PET, is a thermoplastic polymer resin of the polyester family, semi-crystalline, and non-biodegradable. PET can be a transparent, amorphous thermoplastic when rapidly cooled, but when cooled slowly, it acts as a semi-crystalline plastic. PET is produced from the polymerization of ethylene glycol (EG) and terephthalic acid (TPA) [10].
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PET can be processed via typical molding (injection, blown, or extrusion). It is used to fabricate thin layer products in the form of a stretched film, and can also fabricate carbonated beverage bottles because of its strength, toughness, stability, and heat-resistant material.
Despite its convenient use in various products, PET waste is a significant portion of plastic pollution (Kim et al., 2020). The use of plastics has become an intrinsic part of modern society, with annual manufacturing surpassing 350 million metric tons, PET being one of the most extensively used [7]. In 2017, PET production capacity exceeded 30 million tons per annum. 485 billion PET bottles were manufactured in 2016 and 583.3 billion plastic bottles were predicted to be manufactured in 2021 [8]. PET’s exceptional properties of durability, mechanical strength, and transparency cause it to be incorporated in almost every disposable carbonated bottle. But the short lifetime of the large-volume products it is used in has recently brought attention to researchers understanding the serious implications of continued manufacturing of nonbiodegradable compounds. This has sparked scientists to conduct life cycle assessment (LCA) analyses; an LCA conducted by Alfarsi and Primadasa underscored that the manufacturing process of PET contributes to 49 environmental impact categories, including global warming potential, human toxicity, and marine aquatic ecotoxicity (Baldowska-Witos et al., 2021). In previous years, there has been increasing amounts of research done in attempts to degrade or detect PET levels in a given system. However, they have been proven unsuccessful thus far.
A recent finding noted endogenous ADP1 muconate transporter MucK, a member of the major facilitator superfamily, as a potentially more efficient alternative to TpaK, a TPA transporter, at enhancing TPA uptake [2]. We are leveraging this research to develop a biosensor using plasmids to measure TPA production, which can assist in monitoring PET degradation [6]. By building plasmids using various importers like MucK and TpaK, transcription factors, promoters, and sigma factors (subunits of all bacterial RNA polymerases), we can find the most specific and efficient combination to detect TPA and therefore PET as a whole [5]. By integrating sfGFP and mScarlet, different fluorescent proteins, into our plasmids, we will use the level of fluorescence to determine the relative amount of TPA present, thereby determining the extent of PET hydrolysis.
In addition to developing a plasmid biosensor to detect TPA, we remain interested in investigating how similar TpaK and MucK are to each other, at the nucleotide, amino acid, and structural levels. In our Docking efforts, we will translate the sequences of both genes from the plasmids we used, and use AlphaFold to predict the structures. Finally, with Pymol, we can import these structures and assess their overlay. Ultimately, our project, designing a biosensor to detect the TPA, can be used by other scientists in their research to find more effective ways to degrade PET. They can then potentially confirm if their degradation methods are efficient enough by determining how much of the thermoplastic polyester resin has broken down.
[1] Allison Z. Werner (2021, July 12). Tandem Chemical Deconstruction and biological upcycling of poly(ethylene terephthalate) to β-ketoadipic acid by pseudomonas Putida KT2440. Metabolic Engineering. https://www.sciencedirect.com/science/article/pii/S1096717621001154
[2] Beckham, G. T., Bentley, G. J., Choi, K. Y., Franden, M. A., Herrou, J., Jha, R. K., Johnson, C. W., Livak, K. J., Portnoy, V. A., Rorrer, N. A., Sandberg, T. E., Schmidt-Rose, T., … Acero, E. H. (2020, October 1). Gene amplification, laboratory evolution, and Biosensor screening reveal Muck as a terephthalic acid transporter in Acinetobacter Baylyi ADP1. Metabolic Engineering. https://www.sciencedirect.com/science/article/abs/pii/S1096717620301543
[3] Bell, E. L., Smithson, R., Kilbride, S., Foster, J., Hardy, F. J., Ramachandran, S., Tedstone, A. A., Haigh, S. J., Garforth, A. A., Day, P. J. R., Levy, C., Shaver, M. P., & Green, A. P. (2022, August 11). Directed evolution of an efficient and thermostable pet depolymerase. Nature News. https://www.nature.com/articles/s41929-022-00821-3
[4] Carr, C. M., Clarke, D. J., & Dobson, A. D. W. (2020, October 19). Microbial polyethylene terephthalate hydrolases: Current and future perspectives. Frontiers. https://www.frontiersin.org/articles/10.3389/fmicb.2020.571265/full
[5] Heyde SAH;Arnling Bååth J;Westh P;Nørholm MHH;Jensen K; (n.d.). Surface display as a functional screening platform for detecting enzymes active on pet. Microbial cell factories. https://pubmed.ncbi.nlm.nih.gov/33933097/
[6] Meyer, A. J., Segall-Shapiro, T. H., Glassey, E., Zhang, J., & Voigt, C. A. (2018, November 26). Escherichia coli “marionette” strains with 12 highly optimized small-molecule sensors. Nature News. https://www.nature.com/articles/s41589-018-0168-3
[7] Pet global bottle production 2021. Statista. https://www.statista.com/statistics/723191/production-of-polyethylene-terephthalate-bottles-worldwide/
[8] Plastic Insight. Plasticsinsight.com. (n.d.). https://www.plasticsinsight.com/resin-intelligence/resin-prices/polyethylene-terephthalate/Published by Statista Research Department, & 24, M. (2023, March 24).
[9] Sadler, J. C., & Wallace, S. (2021, June 10). Microbial synthesis of vanillin from waste poly(ethylene terephthalate). Green chemistry : an international journal and green chemistry resource : GC. https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8256426/
[10] What is pet plastic?. American Beverage Association. (n.d.). https://www.americanbeverage.org/education-resources/blog/post/what-is-pet/#:~:text=September%2019%2C%202022,and%20made%20to%20be%20remade.